Field of the Invention
The present invention relates to enhancement of bioluminescence signatures of a chemical reaction in an organism or tissue by positioning the organism or tissue near metallic particles.
Background of the Related Art
Metal-Enhanced Fluorescence has been described in detail over the last 5 years as a technology for enhancing fluorescence1-3, phosphorescence4,5 and chemiluminescence6 signatures by the close proximity of metallic nanostructures. In the near-field, i.e. at distances less than 100 Å from the surface, excited states can non-radiatively induce mirror dipoles in the metallic surface, the surface plasmons in turn, radiating the coupled quanta efficiently. Typically, one observes enhanced far-field radiation (lower detection limits when MEF is applied to Immunoassays7,8), with considerably shorter luminescent lifetimes, which are though to reflect the very short plasmon lifetimes themselves.9 Since the lifetimes are considerably reduced, one often observes enhanced luminophore photostability, as the luminescent species spend less time in an excited state and are therefore less prone to destructive excited state process, such as photo-oxidation. Subsequently, MEF affords for ultra-bright and ultra-stable luminescence probes10 and detection platforms7,8,11 to be realized.
For both fluorescent and phosphorescent probes, which generically require an external light source for electronic excitation, an additional electric field effect also enhances the far-field luminescent yield by an increase in the absorption-cross section of the fluorophore in the coupled fluorophore-metal system. For systems where no external light source is used for excitation, such as for chemiluminescence, (chemically induced electronic excited states), dramatic MEF enhancements have also been reported, even in the absence of an electric field component, with ≈1000-fold increases in chemiluminescence reported.6 
Traditionally, bioluminescence signatures are relatively weak as compared to fluorescence-based probes with sensitive detectors often employed.12 Bioluminescence is found in microorganisms [i.e., some bacteria (mostly marine forms, e.g., Vibrio fischeri), fungi, and dinoflagellates], insects (e.g., the firefly, Photinus pyradis), some crustaceans (i.e., Cypridine hilgendorfi), jellyfish, worms and other invertebrates and even in mammals. Although the biochemical mechanism of luminescence is known to vary (i.e., the luminescence system found in bacteria is different from that found in fireflies and dinoflagellates), light production in living organisms is most frequently catalyzed by the enzyme luciferase. Bacterial luciferase is a mixed function oxidase, consisting of two different subunits each with a molecular weight of approximately 40,000 daltons.
Bioluminescence is used for quantitative determinations of specific substances in biology and medicine. For example, luciferase genes have been cloned and exploited as reporter genes in numerous assays, for many purposes. Since the different luciferase systems have different specific requirements, they may be used to detect and quantify a variety of substances. The majority of commercial bioluminescence applications are based on firefly [Photinus pyralis] luciferase.
One of the first and still widely used assays involves the use of firefly luciferase to detect the presence of ATP. It is also used to detect and quantify other substrates or co-factors in the reaction. Any reaction that produces or utilizes NAD(H), NADP(H) or long chain aldehyde, either directly or indirectly, can be coupled to the light-emitting reaction of bacterial luciferase. Another luciferase system that has been used commercially for analytical purposes is the Aequorin system. The purified jellyfish photoprotein, aequorin, is used to detect and quantify intracellular Ca2+ and its changes under various experimental conditions.
These methods suggest activation of the reporter luciferase genes with emission of a readily detectable light signal which allows the monitoring of bacterial response in real-time, by simple luminometry (e.g. fiber optic, luminometers).13,14 The most commonly used systems are the luc gene from the firefly and lux genes from bacterial species of the genus Vibrio. Expression of the lux luciferase operon produces light without any additions, allowing thereby on-line monitoring of gene expression; whereas the expression of firefly luciferase genes requires externally added substrate (luciferin) for luminescence. The bacterial lux system is expressed very effectively in different bacterial strains and this method has been widely applied for different applications.15-17 These luciferases and related reagents are used as reagents for diagnostics, quality control, environmental testing and other such analyses.
However current testing methods are very slow with limited sensitivity and such shortcomings can be critical in regard to bacterial diseases of humans and animals. Thus it would be advantageous to develop a system and method to overcome the shortcomings of prior art testing methods.